This application addresses Support Opportunity for Addiction Research (SOAR) to supplement new investigators who have, or have a commitment of support to conduct research in drug abuse from funding sources other than NIH. Epidemiological studies have shown that there is an increased risk of cardiovascular diseases in children born to women who smoked during pregnancy, but its pathogenesis remains incompletely understood. As one of the major components in cigarette smoking, nicotine is likely to contribute to the fetal programming of cardiovascular disorders. Indeed, our recent studies have demonstrated that nicotine causes development of hypertensive phenotypes in rat offspring. However, the molecular mechanisms underlying nicotine-induced hypertensive phenotypes in offspring are unclear. To understand the mechanisms, two central hypotheses are proposed in this project. The first one is that prenatal nicotine-induced hypertensive responses are mediated by up-regulation of Angiotensin II (Ang II)-mediated signaling pathways, which is being tested in my on-going TRDRP-funded grant project. Given the fact that prenatal nicotine-induced hypertensive responses are associated with vascular Ang II receptor type 2 (AT2R) repression, thus, in this application we will test the following hypothesis that prenatal nicotine exposure causes a gender-specific epigenetic modification of vascular AT2R gene, leading to development of hypertensive phenotypes in offspring. To test this hypothesis, 2 Specific Aims are proposed to determine: 1) whether the AT2R gene repression is associated with a gender- specific AT2R promoter hypermethylation; 2) whether the hypermethylation is associated with an increased reactive oxygen species (ROS) signaling. To achieve these aims, we propose a series of experiments in our established pregnant rat model. Nicotine will be administered to pregnant rats via subcutaneous osmotic minipumps throughout the gestation. The experiments will be conducted at adult offspring. We will measure the vascular AT2R gene expression and its promoter methylation levels, and then determine whether the altered methylation affects transcription factors binding to their putative binding sites at AT2R promoter region. In addition, we will determine whether nicotine-enhanced NADPH oxidase/ROS activity regulates AT2R promoter methylation. We expect that in utero nicotine exposure will cause a gender-dependent epigenetic modification of vascular AT2R expression in association with differential methylation at critical transcription factor binding sites, which may be regulated through nicotine/nAchR-mediated ROS signaling mechanism. The results will provide a novel molecular basis to understand of the fetal programming of adult cardiovascular dysfunction and improve our understanding of tobacco-related fetal programming of adult diseases.